Improved Intrvenous Lentiviral Gene Therapy Based on Endothelial-specific Factor VIII for Hemophilia A

Abstract Background: Hemophilia A (HA) is an X-linked monogenic disorder caused by deficiency of the factor VIII (FVIII, F8) gene in the intrinsic coagulation cascade. The current protein replacement therapy (PRT) of HA has many limitations including short term effectiveness, high cost, and life-time treatment requirement. Gene therapy has become a promising treatment for HA. Orthotopic functional F8 biosynthesis is critical to its coagulation activities. Methods: To investigate targeted F8 expression, we developed a series of advanced lentiviral vectors (LVs) carrying either a universal promoter (EF1α) or a variety of tissue-specific promoters, including endothelial-specific (VEC), endothelial and epithelial-specific (KDR), and megakaryocyte-specific (Gp and ITGA) promoters. Results: To examine tissue specificity, the expression of a B-domain deleted human F8 (F8BDD) gene was tested in human endothelial and megakaryocytic cell lines. Functional assays demonstrated F8 activities of LV-VEC-F8BDD and LV-ITGA-F8BDD in the therapeutic range in transduced endothelial and megakaryocytic cells, respectively. In F8 knockout mice (F8 KO mice F8null mice), intravenous (iv) injection of LVs illustrated different degrees of phenotypic correction as well as anti-F8 immune response for the different vectors. The iv delivery of LV-VEC-F8BDD and LV-Gp-F8BDD achieved 80% and 15% therapeutic F8 activities over 180 days, respectively. Different from the other LV constructs, the LV-VEC-F8BDD displayed a low F8 inhibitory response in the treated F8null mice. Conclusions: The LV-VEC-F8BDD exhibited high LV packaging and delivery efficiencies, with endothelial specificity and low immunogenicity in the F8null mice, thus has a great potential for clinical applications.